Herpes simplex virus is an important human pathogen responsible for a range of diseases from mild uncomplicated mucocutaneous infections to life-threatening ones. Currently, the emergence of Herpes simplex virus resistant strains increased the need for more effective and less cytotoxic drugs for Herpes treatment. In this work, we synthesized a series of oxoquinoline derivatives and experimentally evaluated the antiviral activity against acyclovir resistant HSV-1 strain as well as their cytotoxity profile. The most active compound (3b), named here as Fluoroxaq-3b, showed a promising profile with a better cytotoxicity profile than acyclovir. The theoretical analysis of the structure-activity relationship of these compounds revealed some stereoelectronic properties such as lower LUMO energy and lipophilicity, besides a higher polar surface area and number of hydrogen bond acceptor groups as important parameters for the antiviral activity. Fluoroxaq-3b showed a good oral theoretical bioavailability, according to Lipinski rule of five, with a promising profile for further in vivo analysis.